title: Membrane binding and insertion of the predicted transmembrane domain of human scramblase 1.
abstract: Human phospholipid scramblase 1 (SCR) was originally described as an intrinsic membrane protein catalyzing transbilayer phospholipid transfer in the absence of ATP. More recently, a role as a nuclear transcription factor has been proposed for SCR, either in addition or alternatively to its capacity to facilitate phospholipid flip-flop. Uncertainties exist as well from the structural point of view. A predicted α-helix (aa residues 288-306) located near the C-terminus has been alternatively proposed as a transmembrane domain, or as a protein core structural element. This paper explores the possibilities of the above helical segment as a transmembrane domain. To this aim two peptides were synthesized, one corresponding to the 19 α-helical residues, and one containing both the helix and the subsequent 12-residues constituting the C-end of the protein. The interaction of these peptides with lipid monolayers and bilayers was tested with Langmuir balance surface pressure measurements, proteoliposome reconstitution and analysis, differential scanning calorimetry, tests of bilayer permeability, and fluorescence confocal microscopy. Bilayers of 28 different lipid compositions were examined in which lipid electric charge, bilayer fluidity and lateral heterogeneity (domain formation) were varied. All the results concur in supporting the idea that the 288-306 peptide of SCR becomes membrane inserted in the presence of lipid bilayers. Thus, the data are in agreement with the possibility of SCR as an integral membrane protein, without rejecting alternative cell locations.
authors.affiliation: Unidad de Biofísica (Centro Mixto CSIC, UPV/EHU) and Departamento de Bioquímica, Universidad del País Vasco, P.O. Box 644, 48080 Bilbao, Spain.
authors.forename: Itziar M D
authors.initials: IM
authors.lastname: Posada
authors.forename: Jon V
authors.initials: JV
authors.lastname: Busto
authors.forename: Félix M
authors.initials: FM
authors.lastname: Goñi
authors.forename: Alicia
authors.initials: A
authors.lastname: Alonso
mesh_terms: D000595:Amino Acid Sequence; D002152:Calorimetry, Differential Scanning; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry*; D008560:Membrane Fluidity; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D049294:Phospholipid Transfer Proteins / Q000737:chemistry*; D010743:Phospholipids / Q000737:chemistry*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D055672:Static Electricity
chemical_list: D008051:Lipid Bilayers; C493422:PLSCR1 protein, human; D010455:Peptides; D049294:Phospholipid Transfer Proteins; D010743:Phospholipids
keywords: 1,1,1,3,3,3-hexafluoro-2-propanol; 1,1′-dioctadecyl-3,3,3′,3′-tetramethyl-indodicarbocyanine; 8-aminonaphtalene-1,3,6-trisulfonic acid sodium salt; ANTS; DID; DPX; GUVs; HFIP; LUVs; Lipid phases; MLVs; PC; PE; PG; PI; PLSCR1; PS; SCR; SCR 288–306 peptide; SCR 288–318 peptide; Scramblase; TM; TM19; TM31C; Transbilayer; Transmembrane helix; giant unilamellar vesicles; human phospholipid scramblase 1, or hPLSCR1; large unilamellar vesicles; multilamellar vesicles; p-xylene-bis(pyridinium) bromide; pSM; palmitoyl sphingomyelin; phosphatidylcholine; phosphatidylethanolamine; phosphatidylglycerol; phosphatidylinositol; phosphatidylserine; transmembrane